Hepatotoxicants and carcinogens have been studied for metabolic effect prior to the omic era, and more recently using microarray transcriptomic technology. While the latter approach has greatly expanded knowledge of such compounds, transcriptomic approaches do not actually measure the metabolites (small molecules) and pathways perturbed.
For example, clofibrate is a fibrate type of hypolipidemic drug, and also a hepatotoxicant and carcinogen. It acts on peroxisome proliferator activated receptor alpha (PPARα) receptors. Peroxisome proliferator activated receptors (PPARs) are nuclear hormone receptors that are activated by micromolar concentrations of lipids, fibrates and thiazolidinediones. This subfamily can be divided into three isotypes, designated PPARα, δ, and γ, each with tissue-specific expression. PPARα receptors are particularly abundant in rodents, but are also present in humans. In humans, PPARγ predominates over PPARα, and hepatocyte nuclear factor (HNF) has some similar functions as PPARα in rodents, but both PPAR types are present in rodents and humans. Clofibrate (ethyl-p-chloro-phenoxyisobutyrate; CAS 637-07-0)) is a fibrate type of hypolipidemic (cholesterol lowering) drug, which is also a hepatotoxicant and carcinogen at high levels. It acts predominately on PPARα receptors.
Clofibrates work by activating PPARs, which in turn form heterodimers with retinoid X receptor (RXR), and interact with the peroxisome proliferator response element (PPREs) in gene promoters. PPREs are direct repeats (DR) of a hexanucleotide sequence AGGTCA separated by one nucleotide and are therefore referred to as a DR-1 response element. PPARα and PPARγ play critical roles in the catabolism and storage of fatty acids, whereas the function of PPARδ is less certain. PPARα is the predominant PPAR subtype expressed in liver.
The overall effects of clofibrate are to decrease fat synthesis and increase fat degradation; and to decrease glycolysis and increase gluconeogenesis. In essence, clofibrate mimics the fasted metabolic state. Other effects of clofibrate observed in some studies are: increased oxidative stress; increased cell replication; and increased spontaneous preneoplastic lesions. Short term treatment of clofibrate may not induce transcriptional events as efficiently or at all, as no DNA adducts have been observed. Gonzalez et al. (1998) J Natl Cancer Inst 90: 1702-1709. PPARα regulates genes involved in fatty acid transport, synthesis and oxidation, glucose and lipid metabolism, ketogenesis and Δ5, Δ6, and Δ9-desaturation of fatty acids. Specific genes altered by clofibrate, with possible PPREs are described in Berger et al. (2002) Lipids Health Dis 1: 2 and Hamadeh et al. (2002) Toxicol Sci 67: 219-231.
Clofibrate has been studied at high doses for various durations for its hepatotoxic and carcinogenic effects with microarrays, thus providing a putative map of how clofibrate may affect metabolism. In one study, rats exposed to clofibrate were monitored over time by a combination of histopathology and a transcriptomic approach. After 24 h, there were no microscopic changes to liver after a single exposure of clofibrate or other toxicants. In contrast, after 2 weeks, clofibrate induced hypertrophy. Although a similar set of genes was modified under both conditions, pattern recognition could distinguish the different drug treatments.
These studies demonstrate the predictive biomarker potential of hepatic transcriptomics with respect to liver histopathology changes in response to exposure to hepatotoxicants and carcinogens. Nonetheless, such approaches fail to actually measure the metabolites and pathways perturbed. Thus, there is a need for readily accessible biomarkers of exposure to hepatotoxicants and carcinogens (i.e., biomarkers present in serum, blood, or saliva).